Rhodopsin in the rod outer segment plasma membrane.
Rhodopsin in the rod outer segment plasma membrane.
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DOI：
10.1083/jcb.69.1.29
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发表时间：
1976-04
期刊：
影响因子：
--
通讯作者：
Hall M
中科院分区：
文献类型：
--
作者：
Basinger S;Bok D;Hall M
离体蛙视网膜在体外孵育4小时脉冲的[3H]亮氨酸，然后追逐32小时的非放射性氨基酸混合物。在孵育结束时，从一些视网膜制备光学和电子显微镜放射自显影图。放射自显影显示：（a）在视杆外节的基底盘中的强放射性，（B）在整个视杆外节中均匀分布的扩散标记，和（c）在整个视杆外节质膜中的高浓度标记。在相同条件下孵育，但加入嘌呤霉素，显着抑制所有这些组件的标记。为了鉴定标记的蛋白质，通过SDS圆盘凝胶电泳和凝胶过滤色谱法对来自剩余视网膜的纯化外片段进行生化分析。SDS凝胶电泳显示，约90%的视色素与视杆外段放射性色谱图一致，表明质膜上的放射性标记蛋白是视色素。凝胶过滤层析证明放射性标记的蛋白质与视紫红质而不是视蛋白共层析，并且新合成的视色素是基底盘和质膜两者都以天然构型存在。
Isolated frog retinas were incubated in vitro with a 4-h pulse of [3H]leucine, then chased for 32 h with a nonradioactive amino acid mixture. At the end of the incubation, light and electron microscope autoradiograms were prepared from some of the retinas. The autoradiograms revealed: (a) intense radioactivity in the basal disks of the rod outer segments, (b) diffuse label evenly distributed throughout the rod outer segments, and (c) a high concentration of label in the entire rod outer segment plasma membrane. Incubation under identical conditions, but with puromycin added, significantly inhibited the labeling of all of these components. To identify the labeled proteins, purified outer segments from the remaining retinas were analyzed biochemically by SDS disc gel electrophoresis and gel filtration chromatography. SDS gel electrophoresis showed that about 90% of the total rod outer segment radioactivity chromatographed coincident with visual pigment, suggesting that the radiolabeled protein in the plasma membrane is visual pigment. Gel filtration chromatography demonstrated that the radiolabeled protein co- chromatographed with rhodopsin rather than opsin, and that the newly synthesized visual pigment is both the basal disks and the plasma membrane is present in the native configuration.